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Jean-pierre Cartron - One of the best experts on this subject based on the ideXlab platform.
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The molecular basis of the Rhesus antigen Ew.
Transfusion, 2004Co-Authors: Erwin Strobel, Jean-pierre Cartron, Sabine Hofmann, Matthias F. BauerAbstract:w(ISBT designation 004 011) was first described in 1955. It is defined by a specific antibody, but its molecular Genetic basis has not yet been resolved. STUDY DESIGN AND METHODS: Two individuals serologically characterized to express the rare Rhesus antigen E w were analyzed by sequencing of all 10 exons of the RHCE Gene. RESULTS: A nucleotide exchange at position 500 (T500A) resulting in a Met167Lys amino acid substitution was found in both individuals. Moreover, we show that an individual carrying the E w antigen is capable to produce an alloantibody against the wild-type E antigen. CONCLUSION: The single-point mutation T500A in exon 4 of the RHCE Gene is a molecular basis of the rare Rhesus antigen E w . he Rhesus antigens represent the most complex blood group system on human RBCs, with at least 48 serologic entities described so far. 1 It is
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Evidence that the red cell skeleton protein 4.2 interacts with the Rh membrane complex member CD47.
Blood, 2002Co-Authors: Isabelle Mouro-chanteloup, Jean-pierre Cartron, Pierre Gane, Caroline Le Van Kim, Jean Delaunay, Virginie Nicolas, Mette Johansen, Eric J. Brown, Luanne L. Peters, Yves ColinAbstract:Rhnull red cells are characteristically stomato-spherocytic. This and other evidence suggest that the Rh complex represents a major attachment site between the membrane lipid bilayer and the erythroid skeleton. As an attempt to identify the linking protein(s) between the red cell skeleton and the Rh complex, we analyzed the expression of Rh, RhAG, CD47, LW, and glycophorin B proteins in red cells from patients with hereditary spherocytosis associated with complete protein 4.2 deficiency but normal band 3 (4.2(-)HS). Flow cytometric and immunoblotting analysis revealed a severe reduction of CD47 (up to 80%) and a slower mobility of RhAG on sodium dodecyl sulfate–polyacrylamide gel electrophoresis, possibly reflecting an overglycosylation state. Unexpectedly, 4.2−/− mice, which are anemic, displayed a normal red cell expression of CD47 and RhAG. These results suggest that human protein 4.2, through interaction with CD47, is involved in the skeleton linkage and/or membrane translocation of the Rh complex. However, these potential role(s) of protein 4.2 might be not conserved across species. Finally, the absence or low expression of red cell CD47 in CD47−/− mice and in some humans carrying RHCE Gene variants (D--, D.., and RN), respectively, had no detectable effect on protein 4.2 and RhAG expression. Since these cells are morphologically normal with no sign of hemolysis, it is assumed that CD47 deficiency per se is not responsible for the cell shape abnormalities and for the compensated hemolytic anemia typical of 4.2(-) and Rhnull red cells.
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Two new alleles of the RHCE Gene in Black individuals: the RHCE allele ceMO and the RHCE allele cEMI.
British journal of haematology, 2001Co-Authors: I Mouro, Philippe Rouger, Pierre-yves Le Pennec, Hélène Ansart-pirenne, Geneviève Juszczak, Claude Patereau, M. Verdier, Jérôme Babinet, Michèle Roussel, Jean-pierre CartronAbstract:Six unrelated individuals of Afro-Caribbean origin, whose red cells have a marked reduction of the Rhe antigen expression, have been identified. All exhibited the same serological profile with anti-e monoclonal antibodies and lacked expression of the high frequency e-related antigen hrS. Transcripts and genomic analysis showed that these phenotypes resulted from the presence of two new RHCE alleles, ceMO and cEMI. The ceMO allele corresponded to a RHCE Gene carrying a G667T mutation (exon 5) and was detected at the homozygous state in sample 1 and at the heterozygous state in samples 2–6. The G667T mutation resulted in a Val223Phe substitution on the RHCE polypeptide, in close proximity to Ala226 (e-antigen polymorphism), which might account for the altered expression of e. The ceMO allele is also associated with the lack of expression of the hrS antigen. The absence of the hrS antigen expression may have implications in transfusion as hrS-negative individuals may develop clinically significant antibodies. The cEMI allele corresponded to a silent RHE allele carrying a nine nucleotide deletion within exon 3 and was detected at the heterozygous state in sample 2. This deletion resulted in a shortened polypeptide of 414 residues (instead of 417) that was absent (or severely reduced) at the red cell surface, as the E antigen was undetectable using serology and Western blot analysis with anti-E reagents. In DNA-based polymerase chain reaction genotyping for RHE determination, the cEMI allele provided a false positive result as the cells carrying this allele are serologically phenotyped as E-negative. The incidence of this allele in the Black population is unknown but, as shown already for D genotyping, one must exercise caution when genotyping is performed to detect the e/E polymorphism.
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Molecular Defects of the RHCE Gene in Rh-Deficient Individuals of the Amorph Type
Blood, 1998Co-Authors: Baya Chérif-zahar, Pierre Gane, Giorgio Matassi, Virginie Raynal, Wolfgang Mempel, Carmen Perez, Jean-pierre CartronAbstract:The deficiency of Rh proteins on the red blood cells from individuals of the Rhnull amorph type may be the result of homozygosity for a silent allele at the RH locus. This phenotype is also associated with the lack or reduced expression of glycoproteins (Rh50, CD47, LW, and glycophorin B), which interact with Rh polypeptides to form the multisubunit Rh membrane complex. In this study, we describe two molecular alterations affecting the RHCE Gene in two unrelated Rhnull amorph individuals bearing Rh50 and CD47 normal transcripts. The first type of mutation, located at the donor splice-site in intron 4, induced the activation of two cryptic splice-sites within this intron and one such site in exon 4 that all Generated aberrant transcripts. The second type of mutation affected the coding region and introduced a frameshift and a premature stop codon resulting in a shorter predicted protein (398 v 417 residues), including a completely different C-terminus of 76 amino acids. This suggests that protein folding and/or protein-protein interaction mediated by the C-terminal domain of the Rh proteins may play a role in the routing and/or stability of the Rh membrane complex.
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HeteroGeneity of blood group RhE variants revealed by serological analysis and molecular alteration of the RHCE Gene and transcript
British journal of haematology, 1998Co-Authors: F. Noizat-pirenne, I Mouro, Pierre Gane, Y Okubo, Y Hori, Philippe Rouger, P Y Le Pennec, Jean-pierre CartronAbstract:After testing red cells from 12 RhE variants with a panel of anti-E monoclonal antibodies (MoAbs), four patterns of reactivity were detected indicating that the MoAbs may recognize four distinct E epitopes designated epE1, epE2, epE3 and epE4. The variants were classified into four categories (cat EI to EIV) which carried epE1 and epE2, epE1 and epE4, epE1, epE3 and epE4, and all four epitopes, respectively. Molecular analysis of the transcripts and genomic DNA of the variants from cat EI, EII and EIII displayed three distinct Genetic alterations. Cat EI variants exhibited a point mutation (T500A) in exon 4 of the RHCE Gene that resulted in a Met167Lys substitution in the third extracellular loop of the RHCE protein. Cat EII variant carried a hybrid Gene structure characterized by replacement of exons 1-3 (or 2-3) of the RHCE Gene by their specific counterparts in the RHD Gene. This latter variant was also associated with a weak expression of the RhC antigen. In cat EIII variants there was a partial DNA exchange of exon 5 sequences (nt 697 and 712) between the RHCE and the RHD Genes, Generating a hybrid Rh cE-D-cE protein carrying the Glu233 and Val238 substitutions. The serological and molecular studies of the RhE variants indicated that: (i) the RhE antigen is a mosaic composed of at least four epitopes and proline at position 226 is necessary but not sufficient for the full expression of the E antigen, (ii) the lack of RhE epitope(s) is associated with heterogenous molecular alterations of the RHCE Gene, and (iii) amino-acids located on the third and fourth extracellular loops of the RHCE polypeptide are critical for some RhE epitopes expression.
Marion E. Reid - One of the best experts on this subject based on the ideXlab platform.
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Rh null Syndrome: Identification of a Novel Mutation in RHCE Gene.
Blood, 2004Co-Authors: Marion E. Reid, Karina Antero Ribeiro Rosa, Vivien I. Powell, Christine Lomas-francis, Fernando F. Costa, Servio T. Stinghen, Alexandra M. Watanabe, Edna K. Carboni, Joseani P. Baldon, Maria Marta F. JuckschAbstract:Background : Rh null syndrome, which includes the amorph and regulator types, is a rare Genetic disorder characterized by stomatocytosis and chronic mild hemolytic anemia. The suppression of Rh antigen expression for regulator types is attributed to mutations of the RHAG Gene. The deficiency of Rh proteins on the red blood cells (RBCs) from the rare individuals of the Rh null amorph type may be the result of homozygosity for a silent RHCE allele at the RH locus in which the RHD is absent. We studied a Brazilian family transmitting an amorph Rh null disease Gene on a consanguineous background and identified a novel mutation in RHCE Gene causing the loss of function phenotype. Methods : RBCs from two Rh null sisters (G1 and G2) of the amorph type and from family members were analyzed by serology and flow cytometry with monoclonal antibodies specific for Rh (D, C, c, E and e), and LW antigens, for RhAG and CD47 as well as alloantibodies directed against GPB (S, s and U) antigens. Genomic DNA samples and transcripts were tested by PCR and sequence analysis. Results : RBCs from G1 and G2 did not react with the anti-Rh and anti-LW reagents and reacted weakly with anti-RhAG, anti-CD47, anti-s, and anti-U reagents. RBCs from G1 were S+ and from G2 S-. Molecular analyses showed a deletion of the nucleotide at position 963 in exon 7 of the RHCE Gene [GGG(Glu321→GG)] in addition to a deletion of the RHD Gene. This nucleotide deletion in exon 7 introduced a frameshift after Glu321 and a premature stop codon, resulting in a shorter predicted protein with 359 amino acids, including a new C-terminal sequence that is likely to alter the protein conformation and impair the Rh complex assembly. Conclusion : We describe a novel mutation of an amorph Rh null disease Gene leading to a loss of function phenotype. Our findings reinforce previous studies suggesting that protein folding and/or protein-protein interaction mediated by the C-terminal domain of the Rh proteins plays a role in the stability of the Rh membrane complex.
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A new hybrid RHCE Gene (CeNR) is responsible for expression of a novel antigen
Transfusion, 2004Co-Authors: Connie M. Westhoff, Christine Lomas-francis, Jill R. Storry, Phyllis Walker, Marion E. ReidAbstract:BACKGROUND: The red blood cells (RBCs) of a patient, known to have the probable DC(W)(e)/D-- phenotype, typed as D(W)- and Rh32- but were unexpectedly agglutinated by an anti-D(W)/Rh32 serum. The reactivity suggested that the RBCs carried a novel antigen and that the molecular background of this DC(W)(e)/D-- phenotype might be different from those reported. The purpose of this study was to determine the molecular basis of the Rh phenotype. STUDY DESIGN AND METHODS: Samples were obtained for family studies. Standard hemagglutination methods were used. RH mRNA transcripts were isolated by reverse transcription-polymerase chain reaction and sequenced. RESULTS: Sequence analysis revealed that the probond had three different RH transcripts: a normal RHD and two different hybrid transcripts from the RHCE locus, a RHCE-D hybrid with exon 1 from RHCE associated with the D-- haplotype, and a new RHCE-D hybrid. In this new hybrid, exons 1 to 5 are RHCE-specific and exons 6 to 10 correspond to RHD. The C(W) antigen is also encoded by this hybrid Gene. Family studies confirmed that the new RHCE-D hybrid is linked in cis to conventional RHD. CONCLUSION: A new RHCE-D structure is associated with altered expression of C and e antigens in this family and the Generation of a novel low-prevalence antigen (CENR). (Less)
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16Cys encoded by the RHCE Gene is associated with altered expression of the e antigen and is frequent in the R0 haplotype.
British journal of haematology, 2001Co-Authors: Connie M. Westhoff, Leslie E. Silberstein, Dwane E. Wylie, M. Skavdahl, Marion E. ReidAbstract:Serological observations have suggested that numerous D, many e (especially in Blacks), several E, and rare c variants exist within the Rh blood group system. The molecular basis for expression of many of these variants has been elucidated. This study describes five unrelated Caucasians whose red blood cells reacted with polyclonal anti-e but did not react with some monoclonal anti-e, which suggested that they carried a variant e antigen. Molecular investigation revealed the presence of a 48G-->C change (encoding cysteine instead of tryptophan at amino acid 16) in their RHCE Gene. No other differences were found, which suggests that amino acid residues located in the first transmembrane region can affect expression of the e antigen, whose critical residues are on the predicted fourth external loop of the protein. This polymorphism has not previously been observed because polyclonal anti-e does not distinguish this variant from wild type. This position is polymorphic in RHCE alleles and the presence of the 48C nucleotide is often found in the R0 (Dce) haplotype.
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Rhnull Disease: The Amorph Type Results From a Novel Double Mutation in RHCE Gene on D-Negative Background
Blood, 1998Co-Authors: Cheng-han Huang, Marion E. Reid, Ying Chen, Christine SeidlAbstract:Rhnull disease, which includes the amorph and regulator types, is a rare Genetic disorder characterized by stomatocytosis and chronic hemolytic anemia. We studied here a German family transmitting a putative amorph Rhnull disease Gene and identified a rare mutation causing the loss-of-function phenotype. We analyzed the genomic and transcript structure of RH30, RH50, and CD47, the three loci thought to be most critical for expression of the Rh complex in the red blood cell membrane. We showed that in this family the Rh50 and CD47 transcripts were normal in primary sequence. However, the RH30 locus contained an unusual double mutation in exon 7 of the RHCE Gene, in addition to a deletion of the RhD Gene. The mutation targeted two adjacent codons in multiple arrangements probably via the mechanism of microGene conversion. One scheme entails a noncontiguous deletion of two nucleotides, [ATT(Ile322)-->AT] and [CAC(His323)-->CC], whereas the other involves a T-->C transition [ATT(Ile322)--> ATC] and a dinucleotide deletion [CAC(His323)-->C]. They caused the same shift in open reading frame predicted to encode a shortened protein with 398 amino acids. The loss of two transmembrane domains and gain of a new C-terminal sequence are likely to alter the protein conformation and impair the Rh complex assembly. Our findings establish the molecular identity of an amorph Rhnull disease Gene, showing that Rh30 and Rh50 are both essential for the functioning of the Rh structures as a multisubunit complex in the plasma membrane.
I Mouro - One of the best experts on this subject based on the ideXlab platform.
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Two new alleles of the RHCE Gene in Black individuals: the RHCE allele ceMO and the RHCE allele cEMI.
British journal of haematology, 2001Co-Authors: I Mouro, Philippe Rouger, Pierre-yves Le Pennec, Hélène Ansart-pirenne, Geneviève Juszczak, Claude Patereau, M. Verdier, Jérôme Babinet, Michèle Roussel, Jean-pierre CartronAbstract:Six unrelated individuals of Afro-Caribbean origin, whose red cells have a marked reduction of the Rhe antigen expression, have been identified. All exhibited the same serological profile with anti-e monoclonal antibodies and lacked expression of the high frequency e-related antigen hrS. Transcripts and genomic analysis showed that these phenotypes resulted from the presence of two new RHCE alleles, ceMO and cEMI. The ceMO allele corresponded to a RHCE Gene carrying a G667T mutation (exon 5) and was detected at the homozygous state in sample 1 and at the heterozygous state in samples 2–6. The G667T mutation resulted in a Val223Phe substitution on the RHCE polypeptide, in close proximity to Ala226 (e-antigen polymorphism), which might account for the altered expression of e. The ceMO allele is also associated with the lack of expression of the hrS antigen. The absence of the hrS antigen expression may have implications in transfusion as hrS-negative individuals may develop clinically significant antibodies. The cEMI allele corresponded to a silent RHE allele carrying a nine nucleotide deletion within exon 3 and was detected at the heterozygous state in sample 2. This deletion resulted in a shortened polypeptide of 414 residues (instead of 417) that was absent (or severely reduced) at the red cell surface, as the E antigen was undetectable using serology and Western blot analysis with anti-E reagents. In DNA-based polymerase chain reaction genotyping for RHE determination, the cEMI allele provided a false positive result as the cells carrying this allele are serologically phenotyped as E-negative. The incidence of this allele in the Black population is unknown but, as shown already for D genotyping, one must exercise caution when genotyping is performed to detect the e/E polymorphism.
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HeteroGeneity of blood group RhE variants revealed by serological analysis and molecular alteration of the RHCE Gene and transcript
British journal of haematology, 1998Co-Authors: F. Noizat-pirenne, I Mouro, Pierre Gane, Y Okubo, Y Hori, Philippe Rouger, P Y Le Pennec, Jean-pierre CartronAbstract:After testing red cells from 12 RhE variants with a panel of anti-E monoclonal antibodies (MoAbs), four patterns of reactivity were detected indicating that the MoAbs may recognize four distinct E epitopes designated epE1, epE2, epE3 and epE4. The variants were classified into four categories (cat EI to EIV) which carried epE1 and epE2, epE1 and epE4, epE1, epE3 and epE4, and all four epitopes, respectively. Molecular analysis of the transcripts and genomic DNA of the variants from cat EI, EII and EIII displayed three distinct Genetic alterations. Cat EI variants exhibited a point mutation (T500A) in exon 4 of the RHCE Gene that resulted in a Met167Lys substitution in the third extracellular loop of the RHCE protein. Cat EII variant carried a hybrid Gene structure characterized by replacement of exons 1-3 (or 2-3) of the RHCE Gene by their specific counterparts in the RHD Gene. This latter variant was also associated with a weak expression of the RhC antigen. In cat EIII variants there was a partial DNA exchange of exon 5 sequences (nt 697 and 712) between the RHCE and the RHD Genes, Generating a hybrid Rh cE-D-cE protein carrying the Glu233 and Val238 substitutions. The serological and molecular studies of the RhE variants indicated that: (i) the RhE antigen is a mosaic composed of at least four epitopes and proline at position 226 is necessary but not sufficient for the full expression of the E antigen, (ii) the lack of RhE epitope(s) is associated with heterogenous molecular alterations of the RHCE Gene, and (iii) amino-acids located on the third and fourth extracellular loops of the RHCE polypeptide are critical for some RhE epitopes expression.
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Characterization of the recombination hot spot involved in the genomic rearrangement leading to the hybrid D-CE-D Gene in the D(VI) phenotype.
American journal of human genetics, 1997Co-Authors: Giorgio Matassi, I Mouro, B Chérif-zahar, J.-p. CartronAbstract:In the Caucasian population, the RH locus of RhD-positive individuals is composed of two homologous Genes, RHD and RHCE, arranged in tandem but of a single Gene, RHCE, in RhD-negative individuals. Many variants recently characterized carry rearranged RH Genes, most often by an unidirectional segmental DNA-exchange (Gene-conversion) event. In D(VI) variants of type II, RHD is a D-CE-D hybrid Gene in which the DNA fragment carrying exons 4-6 has been replaced by the corresponding sequences from the RHCE Gene. To identify precisely and characterize the two transition sites, we have studied, by both PCR and sequence analysis, a genomic region between the 3' end of intron 3 and exon 7 in normal RHCE and RHD Genes as well as in D(VI) DNA. We show that the D-CE breakpoint is located in intron 3, within a 250-bp fragment comprising an Alu S sequence, and that the CE-D breakpoint lies within a 39-bp fragment in intron 6. This Alu S sequence (and the 100-bp region immediately downstream) most likely defines a recombination hot spot, since there lies also the 5' breakpoint of different rearrangement events leading to D-CE and CE-D transitions in hybrid D(VI),DFR and Dc-,R(N) Gene complexes, respectively.
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Molecular analysis of blood group Rh transcripts from a rGr variant
British journal of haematology, 1996Co-Authors: I Mouro, Jean-pierre Cartron, Pierre Gane, Yves Colin, Emmanuel Collec, Teresa Zelinski, Caroline Le Van KimAbstract:The Rh blood group antigens D, Cc and Ee are encoded by two related Genes, RHD and RHCE. The RhG antigen (Rh12) is associated with the expression of RhC and/or RhD, except in rare variant red cells. Here we have determined the molecular basis of G expression in the absence of D and C in the r G r phenotype. Nucleotide sequence analysis revealed that the r G allele resulted either from a segmental DNA exchange between part of exon 2 of the RHCE Gene and the equivalent region of the RHCE or RHD Genes or from a crossing over between positions nt150 and nt178 of the RHCE and RHCE Genes. The predicted protein encoded by the hybrid r G Gene (c-C-e or c-D-e) carries Ile60, Ser68 and Ser103 (as C and D polypeptides) ; any of these positions appear to be critical in the formation of the G antigen. In addition, Cys16 was found to be important in the phenotypic expression of C.
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Molecular Basis of the RhCW (Rh8) and RhCX (Rh9) Blood Group Specificities
Blood, 1995Co-Authors: I Mouro, P Y Le Pennec, Yves Colin, P Sistonen, J.-p. Cartron, C. Le Van KimAbstract:The Rh blood group antigens are encoded by two highly related Genes, RHD and RHCE, and the sequence of the common alleles (D, Ce, CE, ce, and cE) of these Genes has been previously elucidated. In this report, Rh transcripts and Gene fragments have been amplified using polymerase chain reaction from the blood of donors with the CW+ andCX+ phenotypes. Sequence analysis indicated that the expression of the CW (Rh8) and CX (Rh9) antigens are associated with point mutations in the RHCE Gene, which provides the definitive evidence that the CW and CX specificities are encoded by the same Gene as the Cc and Ee antigens. As compared with the common (CW- and CX-) transcripts of the RHCE Gene, the CW+ and CX+ cDNAs exhibited A122G and G106A transitions that resulted in Gln41Arg and Ala36Thr amino acid substitutions in the CW+ and CX+ polypeptides, respectively. Therefore, although the CW and CX specificities behave serologically as if they were allelic, they cannot not be considered, stricto sensu, as the products of antithetical allelic forms of the RHCE Gene. Based on the CW-/CW+ nucleotide polymorphism, a polymerase chain reaction assay useful for diagnosis purposes has been developed that detects the presence of the CW+ allele by the use of an allele-specific primer.
Li Xiuzhen - One of the best experts on this subject based on the ideXlab platform.
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Polymorphism of RhD Gene in RhD-negative blood donors in Henan Province
Journa of Henan Medical University, 2002Co-Authors: Li XiuzhenAbstract:Aim: To explore the characteristics of RhD Gene structure among RhD negative unrelated blood donors in Henan Province. Methods:PCR SSP was used to amplify RhD Gene exons 2,3,4,5,6,7,9,10 and intron 4 as well as RHCE Gene intron 4 in 80 RhD negative blood donors which were determined by routine serological method. Results:Out of 80 RhD negative blood donors, 53(66.2%) cases were complete RhD Gene deletion,7(8.7%) cases were partial RhD Gene deletion, of which most were RhD Gene exon 3～9 deletion and 21 cases carried the intact RhD Gene(25%). Conclusions: The results suggest that there are multiple Genetic mechanisms to Generate RhD negative trait for people in Henan Province. To identify Rh types by using DNA analysis in white people is not fit for Chinese people.It is necessary to develop clinical testing method suitable for Chinese pelple.
Michael C. Snabes - One of the best experts on this subject based on the ideXlab platform.
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Single-cell analysis of the RhD blood type for use in preimplantation diagnosis in the prevention of severe hemolytic disease of the newborn☆
American Journal of Obstetrics and Gynecology, 1995Co-Authors: Ignatia B. Van Den Veyver, Jean-pierre Cartron, Yves Colin, Samuel S. Chong, Juan Cota, Phillip R. Bennett, Nicholas M. Fisk, Alan H. Handyside, Caroline Le Van Kim, Michael C. SnabesAbstract:Abstract OBJECTIVE: Our purpose was to develop a molecular assay to determine the fetal RhD blood type on single diploid cells, including blastomeres. STUDY DESIGN: Polymerase chain reaction amplification of a 99 bp deoxyribonucleic acid fragment of the RhD Gene or a 113 bp fragment from the RHCE Gene was performed from 20 venous blood samples and 20 amniotic fluid samples and from 60 single-cultured lymphoblasts and 12 media blanks mixed in a blinded fashion. This reaction was similarly tested after whole-genome amplification on 10 lymphoblasts and seven human blastomeres. RESULTS: Deoxyribonucleic acid amplification was successful and correct from all genomic deoxyribonucleic acid samples. Ninety-seven percent of single cells amplified: correct diagnosis was made in 96%. Five blastomeres successfully amplified. No media blanks produced amplified, contaminating deoxyribonucleic acid. CONCLUSIONS: The RhD blood type can be determined reliably from single cells and can be used for preimplantation Genetic diagnosis for the prevention of rhesus hemolytic disease.
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Single-cell analysis of the RhD blood type for use in preimplantation diagnosis in the prevention of severe hemolytic disease of the newborn.
American journal of obstetrics and gynecology, 1995Co-Authors: Ignatia B. Van Den Veyver, Y. Colin, J.-p. Cartron, C. Le Van Kim, Samuel S. Chong, Juan Cota, Phillip R. Bennett, Nicholas M. Fisk, Alan H. Handyside, Michael C. SnabesAbstract:Our purpose was to develop a molecular assay to determine the fetal RhD blood type on single diploid cells, including blastomeres. Polymerase chain reaction amplification of a 99 bp deoxyribonucleic acid fragment of the RhD Gene or a 113 bp fragment from the RHCE Gene was performed from 20 venous blood samples and 20 amniotic fluid samples and from 60 single-cultured lymphoblasts and 12 media blanks mixed in a blinded fashion. This reaction was similarly tested after whole-genome amplification on 10 lymphoblasts and seven human blastomeres. Deoxyribonucleic acid amplification was successful and correct from all genomic deoxyribonucleic acid samples. Ninety-seven percent of single cells amplified; correct diagnosis was made in 96%. Five blastomeres successfully amplified. No media blanks produced amplified, contaminating deoxyribonucleic acid. The RhD blood type can be determined reliably from single cells and can be used for preimplantation Genetic diagnosis for the prevention of rhesus hemolytic disease.